Summary: The biochemical mechanism by which botulinum neurotoxins enter and act upon nerve cells is largely unkown. Part of the entry process involves unfolding the toxin protein followed by the translocation of this linear form through a channel leading to the cellular interior. We did the first direct study of this process using BoNT type A and E on cultured spinal cord neurons. We find that internalization of each toxin has a unique pH requirement. BoNT/A requires a greater degree of acidification than does BoNT/E. This approach is being expanded to include the study of BoNT/B and tetanus neurotoxin. The aim is to do the first long-term recovery study for these two toxins, correlate their activity with neurotransmitter release and examine the pH requirement for each.